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Neuroprotective Effect of HIF Prolyl Hydroxylase Inhibition in an In Vitro Hypoxia Model.

ABSTRACT: A novel potent analog of the branched tail oxyquinoline group of hypoxia-inducible factor (HIF) prolyl hydroxylase inhibitors, neuradapt, has been studied in two treatment regimes in an in vitro hypoxia model on murine primary hippocampal cultures. Neuradapt activates the expression of HIF1 and HIF2 target genes and shows no toxicity up to 20 ?M, which is more than an order of magnitude higher than its biologically active concentration. Cell viability, functional activity, and network connectivity between the elements of neuronal networks have been studied using a pairwise correlation analysis of the intracellular calcium fluctuations in the individual cells. An immediate treatment with 1 ?? and 15 ?? neuradapt right at the onset of hypoxia not only protects from the death, but also maintains the spontaneous calcium activity in nervous cells at the level of the intact cultures. A similar neuroprotective effect in the post-treatment scenario is observed for 15 ??, but not for 1 ?? neuradapt. Network connectivity is better preserved with immediate treatment using 1 ?? neuradapt than with 15 ??, which is still beneficial. Post-treatment with neuradapt did not restore the network connectivity despite the observation that neuradapt significantly increased cell viability at 1 ?? and functional activity at 15 ??. The preservation of cell viability and functional activity makes neuradapt promising for further studies in a post-treatment scenario, since it can be combined with other drugs and treatments restoring the network connectivity of functionally competent cells.

SUBMITTER: Savyuk M

PROVIDER: S-EPMC7463909 | biostudies-literature | 2020 Jul

REPOSITORIES: biostudies-literature

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Neuroprotective Effect of HIF Prolyl Hydroxylase Inhibition in an In Vitro Hypoxia Model.

Savyuk Maria M Krivonosov Mikhail M Mishchenko Tatiana T Gazaryan Irina I Ivanchenko Mikhail M Khristichenko Anna A Poloznikov Andrey A Hushpulian Dmitry D Nikulin Sergey S Tonevitsky Evgeny E Abuzarova Guzal G Mitroshina Elena E Vedunova Maria M

Antioxidants (Basel, Switzerland) 20200724 8

A novel potent analog of the branched tail oxyquinoline group of hypoxia-inducible factor (HIF) prolyl hydroxylase inhibitors, neuradapt, has been studied in two treatment regimes in an in vitro hypoxia model on murine primary hippocampal cultures. Neuradapt activates the expression of HIF1 and HIF2 target genes and shows no toxicity up to 20 μM, which is more than an order of magnitude higher than its biologically active concentration. Cell viability, functional activity, and network connectivi ...[more]

PMID: 32722310

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Project description:BACKGROUND:In muscular dystrophy and old age, skeletal muscle repair is compromised leading to fibrosis and fatty tissue accumulation. Therefore, therapies that protect skeletal muscle or enhance repair would be valuable medical treatments. Hypoxia-inducible factors (HIFs) regulate gene transcription under conditions of low oxygen, and HIF target genes EPO and VEGF have been associated with muscle protection and repair. We tested the importance of HIF activation following skeletal muscle injury, in both a murine model and human volunteers, using prolyl hydroxylase inhibitors that stabilize and activate HIF. METHODS:Using a mouse eccentric limb injury model, we characterized the protective effects of prolyl hydroxylase inhibitor, GSK1120360A. We then extended these studies to examine the impact of EPO modulation and infiltrating immune cell populations on muscle protection. Finally, we extended this study with an experimental medicine approach using eccentric arm exercise in untrained volunteers to measure the muscle-protective effects of a clinical prolyl hydroxylase inhibitor, daprodustat. RESULTS:GSK1120360A dramatically prevented functional deficits and histological damage, while accelerating recovery after eccentric limb injury in mice. Surprisingly, this effect was independent of EPO, but required myeloid HIF1?-mediated iNOS activity. Treatment of healthy human volunteers with high-dose daprodustat reduced accumulation of circulating damage markers following eccentric arm exercise, although we did not observe any diminution of functional deficits with compound treatment. CONCLUSION:The results of these experiments highlight a novel skeletal muscle protective effect of prolyl hydroxylase inhibition via HIF-mediated expression of iNOS in macrophages. Partial recapitulation of these findings in healthy volunteers suggests elements of consistent pharmacology compared to responses in mice although there are clear differences between these two systems.

Project description:Introduction: We previously reported that malignant hypertension is associated with impaired capillary density of target organs. Here, we tested the hypothesis that stabilization of hypoxia-inducible factor (HIF) in a modified "preconditioning" approach prevents the development of malignant hypertension. To stabilize HIF, we employed pharmacological inhibition of HIF prolyl hydroxylases (PHD), that profoundly affect HIF metabolism. Methods: Two-kidney, one-clip renovascular hypertension (2K1C) was induced in rats; controls were sham operated. 2K1C rats received either intermittent injections of the PHD inhibitor ICA (2-(1-chloro-4-hydroxyisoquinoline-3-carboxamido) acetate) or placebo. Thirty-five days after clipping, the frequency of malignant hypertension was assessed (based on weight loss and the occurrence of characteristic vascular lesions). In addition, kidney injury was compared between all ICA treated versus all placebo treated 2K1C, regardless of the occurrence of malignant hypertension. HIF stabilization was evaluated by immunohistochemistry, and HIF target gene expression by RT-PCR. Results: Blood pressure was elevated to the same degree in ICA- and placebo-treated 2K1C compared to control rats. ICA treatment did not affect the frequency of malignant hypertension or the extent of kidney tissue fibrosis, inflammation, or capillary density. There was a trend towards higher mortality and worse kidney function in ICA-treated 2K1C rats. ICA increased the number of HIF-1α-positive renal tubular cell nuclei and induced several HIF-1 target genes. In contrast, expression of HIF-2α protein as well as HIF-2 target genes were markedly enhanced by 2K1C hypertension, irrespective of ICA treatment. Discussion: We conclude that intermittent PHD inhibition did not ameliorate severe renovascular hypertension in rats. We speculate that the unexpected strong renal accumulation of HIF-2α in renovascular hypertension, which could not be further augmented by ICA, may contribute to the lack of a benefit from PHD inhibition.

Dataset Information

Neuroprotective Effect of HIF Prolyl Hydroxylase Inhibition in an In Vitro Hypoxia Model.

Publications

Neuroprotective Effect of HIF Prolyl Hydroxylase Inhibition in an In Vitro Hypoxia Model.

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